Molecular cloning of the cancer metastasis associated gene and its clinical applications.

癌症转移相关基因的分子克隆及其临床应用。

基本信息

批准号：
10671117
负责人：
YOSHIDA Kazuhiro
金额：
$ 2.05万
依托单位：
Hiroshima University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 1999
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10671117/
关键词：
Cancer metastasis Metastasis Associated gene cDNA differential display

项目摘要

We have established two human cell lines which cause metastasis written below.1) One clone was established by transfecting with DNA from highly metastatic melanoma cells (A375 melanoma cell) into non-metastatic mouse cell line (AH8 control). This cells highly metastasize to lung when transplanted to the nude mouse.2) Human breast cancer cell line, MDA 435, was transplanted repeatedly to nude mice and metastatic clones to the lung, MDA 4A4 and non-metastatic clone, MDA 2C5 were established.We have previously isolated 4 clones by cDNA differential display method and these clones were revealed as new molecules after homology search by the gene bank. The sizes of these clones by Northern blot were about 2.5kb and 4.5kb as a single band, respectedly. The expressions of the clones were detected only in 4A4 clones and not detected in 2C5 suggesting that these molecules are closely related to cancer metastasis.We next transplanted 4A4 and 2C5 cells into nude mice and extracted RNA from each cells and Northern blot analysis was performed. Interestingly, these clones were not necessarily highly expressed in metastatic cells suggesting that the expression levels of these molecules changes according to the conditions of the circumstances.Moreover, we have tried to detect the mRNA expression profiles of 4A4 and 2C5 clones by DNA microarray system using 20000 human mRNA clones and now more than 200 clones are listed as candidate molecule as metastasis related gene and close analysis including the 4 DNA clones which were isolated previously, are on going now.

我们已经建立了两种引起转移的人类细胞系，如下所示。1）通过将高度转移性黑色素瘤细胞（A375黑色素瘤细胞）的DNA转染至非转移性小鼠细胞系（AH8对照）中，建立了一种克隆。该细胞移植到裸鼠后高度转移至肺。2)将人乳腺癌细胞系MDA 435反复移植到裸鼠体内，建立肺转移克隆MDA 4A4和非转移克隆MDA 2C5。我们之前通过cDNA差异展示方法分离了4个克隆，这些克隆经过基因库同源搜索后被揭示为新分子。通过 Northern blot 检测，这些克隆的单条带大小分别约为 2.5kb 和 4.5kb。这些克隆的表达仅在4A4克隆中检测到，而在2C5中未检测到，表明这些分子与癌症转移密切相关。接下来，我们将4A4和2C5细胞移植到裸鼠体内，并从每个细胞中提取RNA并进行Northern印迹分析。有趣的是，这些克隆不一定在转移细胞中高表达，这表明这些分子的表达水平根据环境条件而变化。此外，我们尝试使用DNA微阵列系统检测4A4和2C5克隆的mRNA表达谱。 20000个人类mRNA克隆和现在超过200个克隆被列为转移相关基因的候选分子，并且包括之前分离的4个DNA克隆在内的密切分析正在进行中。